A functional Campylobacter jejuni maf4 gene results in novel glycoforms on flagellin and altered autoagglutination behaviour.
Flagellin of Campylobacter jejuni is extensively modified with (derivatives of) pseudaminic acid. The flagellar glycosylation locus contains several genes with homopolymeric G-tracts prone to slipped-strand mispairing, some of which belong to the maf gene family. We investigated the function of the putative phase-variable maf4 gene of C. jejuni strain 108. A constructed maf4 mutant displayed unaltered flagella assembly and bacterial motility. 2D-PAGE analysis revealed that the flagellin of strain 108 migrated at a more acidic pI than the protein of the Maf4 mutant. MS-MS in combination with high-resolution matrix-assisted laser desorption/ionization Fourier transform ion cyclotron MS (MALDI-FT-ICR-MS) on flagellin-derived glycopeptides showed that the flagellins of the mutant lacked two previously unidentified modifications of pseudaminic acid. These glycoforms carried additional CO(2) and C(2)H(2)O(2) groups, consistent with the more acidic pI of the wild-type flagellin. Phenotypically, the maf4 mutant displayed strongly delayed bacterial autoagglutination. Collectively, our results suggest that the presence of a functional Maf4 expands the flagellin glycan repertoire with novel glycoforms of pseudaminic acid and, in the event of phase variation, alters the population behaviour of C. jejuni.
van Alphen
Lieke B
LB
Department of Infectious Diseases and Immunology, Utrecht University, Yalelaan 1, 3584 CL Utrecht, The Netherlands.
Wuhrer
Manfred
M
Biomolecular Mass Spectrometry Unit, Department of Parasitology, Leiden University Medical Center, Albinusdreef 2, 2300 RC Leiden, The Netherlands.
Bleumink-Pluym
Nancy M C
NMC
Department of Infectious Diseases and Immunology, Utrecht University, Yalelaan 1, 3584 CL Utrecht, The Netherlands.
Hensbergen
Paul J
PJ
Biomolecular Mass Spectrometry Unit, Department of Parasitology, Leiden University Medical Center, Albinusdreef 2, 2300 RC Leiden, The Netherlands.
Deelder
André M
AM
Biomolecular Mass Spectrometry Unit, Department of Parasitology, Leiden University Medical Center, Albinusdreef 2, 2300 RC Leiden, The Netherlands.
van Putten
Jos P M
JPM
Department of Infectious Diseases and Immunology, Utrecht University, Yalelaan 1, 3584 CL Utrecht, The Netherlands.
D000595:Amino Acid Sequence; D001422:Bacterial Adhesion; D001426:Bacterial Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D016123:Campylobacter jejuni / Q000737:chemistry / Q000145:classification / Q000235:genetics / Q000502:physiology*; D002460:Cell Line; D005407:Flagella / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D005408:Flagellin / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D006031:Glycosylation; D006801:Humans; D008969:Molecular Sequence Data; D010802:Phylogeny; D016415:Sequence Alignment
D001426:Bacterial Proteins; D005408:Flagellin